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Analysis of Chromosomal DNA Fragmentation in Apoptosis by Pulsed-Field Gel Electrophoresis

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DNA Electrophoresis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2119))

Abstract

Double-strand DNA break (DSB) formation is a key feature of apoptosis called chromosomal DNA fragmentation. However, some apoptosis inducers introduce DNA damage-induced DSBs prior to induction of apoptotic chromosomal DNA fragmentation. To analyze these distinct breaks, we have developed a method using pulsed-field gel electrophoresis (PFGE) with a rotating gel electrophoresis system (RGE) that enables us to distinguish between apoptotic DSBs and DNA damaging agent-induced DSBs based on their mobility in the electrophoresis gel. Apoptotic DSBs appear as smeared low-molecular weight bands (less than 500 kb), while damage-induced DSBs result in a compact single band (more than 500 kb). Furthermore, using a caspase inhibitor, Z-VAD-FMK, we can confirm whether broken DNA fragments are produced as part of an apoptotic response. Overall, we succeeded in characterizing two individual apoptosis inducers and showed the different effects of those compounds on the induction of DNA breaks.

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Acknowledgments

We would like to thank Editage (www.editage.jp) for English language editing. This research was supported by a Grant-in-Aid for the Cooperative Research Project from Institute of Natural Medicine, University of Toyama in 2014 and by JSPS KAKENHI Grant Number 16 K07119 to T. T. and 17 K08339 to T. I.

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Authors and Affiliations

  1. Department of Pharmacology, Faculty of Medicine, Oita University, Yufu, Oita, Japan

    Takeshi Terabayashi, Asako Tokumaru & Toshimasa Ishizaki

  2. Clinical Engineering Research Center, Faculty of Medicine, Oita University, Yufu, Oita, Japan

    Katsuhiro Hanada

Authors
  1. Takeshi Terabayashi
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  2. Asako Tokumaru
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  3. Toshimasa Ishizaki
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  4. Katsuhiro Hanada
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Corresponding authors

Correspondence to Takeshi Terabayashi or Katsuhiro Hanada .

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Editors and Affiliations

  1. Clinical Engineering Research Center, Faculty of Medicine, Oita University, Yufu, Oita, Japan

    Katsuhiro Hanada

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Terabayashi, T., Tokumaru, A., Ishizaki, T., Hanada, K. (2020). Analysis of Chromosomal DNA Fragmentation in Apoptosis by Pulsed-Field Gel Electrophoresis. In: Hanada, K. (eds) DNA Electrophoresis. Methods in Molecular Biology, vol 2119. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0323-9_8

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  • DOI: https://doi.org/10.1007/978-1-0716-0323-9_8

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0322-2

  • Online ISBN: 978-1-0716-0323-9

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